鱼腥草挥发油和甲基正壬酮对LPS-TLR4/MD-2-TNF-α炎症通路的影响

Effects of Volatile Oil and 2-Undecanone from Houttuynia Cordata Thunb. on LPS-TLR4 /MD-2-TNF-α Inflammation Signaling Pathway

目的探讨鱼腥草挥发油及其单体甲基正壬酮对LPS-TLR4/MD-2-TNF-α炎症通路的影响。方法以TLR4/MD-2阻断剂封闭TLR4/MD-2位点,分别运用蛋白免疫印迹法（Western blot）和酶联免疫吸附测定法（ELISA）对鱼腥草挥发油和甲基正壬酮干预后细胞中TLR4蛋白和炎症因子肿瘤坏死因子α（TNF-α）、白细胞介素1β（IL-1β）、白细胞介素10（IL-10）的表达进行检测。同时建立二甲苯致小鼠耳肿胀炎症模型,对鱼腥草挥发油和甲基正壬酮的体内抗炎药效进行比较。结果在1～10μg·mL^-1浓度范围内短时干预时,鱼腥草挥发油对LPS诱导的RAW264.7细胞中TLR4蛋白的抑制作用优于甲基正壬酮,对炎症因子的作用与甲基正壬酮有一定差异。LPS＋TLR4/MD-2阻断剂＋药物干预组与LPS＋TLR4/MD-2阻断剂组比较,以挥发油干预后细胞中TLR4蛋白的表达差异无统计学意义（P〉0.05）,而以甲基正壬酮干预后细胞中TLR4蛋白的表达明显减少。表明鱼腥草挥发油通过LPS-TLR4/MD-2-TNF-α通路发挥其抗炎作用,而甲基正壬酮可能通过其他途径发挥其抗炎作用。在相同剂量下鱼腥草挥发油的体内抗炎活性优于甲基正壬酮。结论鱼腥草挥发油与甲基正壬酮存在抗炎作用及机制的差异性,应是挥发油中多成分协同作用的结果。

Objective To explore the effects of volatile oil and 2-undecanone from Houttuynia Cordata Thunb.（ H.cordata） on LPS-TLR4 /MD-2-TNF-α signaling pathway. Methods TLR4 /MD-2 blocking agent was used to mask the TLR4 /MD-2 site,then protein expression levels of TLR4 in cells treated with volatile oil and 2-undecanone were analyzed by western blot. ELISA was used to detect the secretion of the inflammatory cytokines such as TNF-α,IL-1β and IL-10. Comparison analysis was then performed from the results of cell experiments in vitro and anti-inflammatory effects through xylene-induced ear edema test in vivo. Results In concentrations between 1 to 10 μg · mL^-1,Houttuynia volatile oil showed better effect than 2-undecanone on inhibition of TLR4 protein in LPS-induced RAW264. 7 cells,and had some differences in the effects on inflammatory factors. Compared with the LPS ＋ TLR4 /MD-2 group,the LPS ＋ TLR4 /MD-2 ＋ volatile oil group had no significant difference in the expression of TLR4 protein（ P〉 0. 05）,but the LPS ＋ TLR4 /MD-2 ＋ 2-undecanone group reduced the expression of TLR4 protein obviously. It appeared that volatile oil exerts its anti-inflammatory effect through LPS-TLR4 /MD-2-TNF-α pathway,but 2-undecanone may exert its anti-inflammatory effect by other means. Houttuynia volatile oil showed better anti-inflammatory activity than 2-undecanone in vivo at the same dose. Conclusion There are some differences in antiinflammatory effects and related mechanisms between volatile oil and 2-undecanone,probably owing to the synergistic effects of multi-ingredients in the volatile oil.